Therapeutic Effects And Mechanism Of Methylene Blue On EAE Mice

Objective:To investigate the effects of methylene blue(MB)on experimtal autoimmune encephalomyelitis(EAE)mice,which is induced by myelin oligodendrocyte glycoprotein(MOG).The proportion of Th17 and Treg cells in the spleen tissue of mice was observed,and the therapeutic effects of MB on EAE mice as well as the possible immunologic mechanism were discussed.Methods:1 24 C57BL/6 mice were randomly divided into three groups:EAE group,MB group and control group,8 mice each group.Then EAE model was established.EAE group and MB group were immunized with MOG35-55 as an antigen,supplemented with pertussis toxin,complete Freund's adjuvant and tuberculin,while control group was not immunized.The day EAE model established were recorded as Day 0.The mice in MB group were given intraperitoneal injection of MB 20mg/kg daily from Day 1.The mice in control group and EAE group were given equal volume of normal saline every day.The onset time and daily neurological scores of mice were observed.2 18 C57BL/6 mice were randomly divided into three groups:EAE group,MB group and control group,6 mice each group.Then EAE model was established.EAE group and MB group were immunized with MOG35-55 as an antigen,supplemented with pertussis toxin,complete Freund's adjuvant and tuberculin,while control group was not immunized.The day EAE model established were recorded as Day 0.The mice in MB group were given intraperitoneal injection of MB 20mg/kg daily from Day 1.The mice in control group and EAE group were given equal volume of normal saline every day.Mice were sacrificed 25 days after immunization(peak phase).The spleens of the mice were removed under sterile conditions.The proportion of Th17 and Treg cells in spleens was tested by Flow Cytometry.Results:1 All mice in MB and EAE group developed EAE,the peak phase is about 25 days after immunization.Compared with EAE group,the neurological scores of MB group was significantly lower(P<0.01),while the onset time of MB group was significantly later(P<0.05).The mice in control group did not develop the disease;2 The proportion of Treg cells in the spleen tissue of each group was significantly different on peak phase.Compared with control group,the proportion of Treg cells in spleen tissue of EAE group was significantly decreased(P<0.01).The proportion of Treg cells in MB group was significantly higher than that in EAE group and control group(P<0.01);3Compared with control group,the percentage of Th17 cells in the spleen tissue of EAE mice was significantly increased(P<0.05).Compared with the EAE group,the percentage of Th17 cells in the spleen tissue was significantly decreased in MB group.(P <0.05).However,there was no significant difference in Th17 cells between MB group and control group(P> 0.05).Conclusions:1 MB intervention can reduce the degree of neurological deficits and delay the onset time of EAE mice.2 MB intervention may reduce the degree of neurological deficits and delay the onset time of EAE mice by its influnence on Th17/Treg cells in the spleen,but the specific mechanism remains to be further studied.